Table 1 Summary of the current published protocols for vascularized hPSC-derived organoids

Pancreas*

HUVEC

Coculture of HUVEC, MSCs and hPSC-derived PP progenitors.

MSC-driven condensation on Matrigel.

• Endothelial networks

• IF: CD31

• In vivo: Anastomosis of human CD31⁺ networks with mouse blood vessels

+

+

Initial % EC: 36%

In vivo reperfusion and non-leaky vessels

[14, 63]

HUVEC

Coculture of HUVEC and hPSC-derived PP progenitors on a hydrogel (“amikagel”).

• IF: vWF

+

–

Initial % EC: 50%

Final % EC: 45–50%

[65]

Liver**

HUVEC

Coculture of HUVEC, MSCs and hPSC-derived hepatic progenitors.

MSC-driven condensation on Matrigel.

• Endothelial networks

• IF: CD31

• In vivo: Anastomosis of human CD31⁺ networks with mouse blood vessels

+

+

Failure of non-vascularized (in vitro) organoids to engraft.

Initial % EC: 11%

[67, 68]

hPSC-EC

Codifferentiation of hPSC into hepatobiliary and endothelial lineage on Matrigel.

• Endothelial networks

• IF and FACS: CD34, CD31

• IF: CD144, CD146

• mRNA expression: JAG1, NOTCH2, HES1

–

–

[71]

HAMEC

EB differentiation of hepatic cells in the presence of HAMEC.

• Endothelial rosettes

• IF: CD31⁺

• mRNA expression: Factor VIII, vWF and other coagulation cascade and fibrinolysis genes

• In vivo: Human CD31⁺ in the rat spleen without migration to the liver.

+

–

Only vascularized organoids supported a sustained production of albumin in vivo up to 14 days after transplantation.

Initial % EC: 30%

Initial % EC: 15%

[70]

HUVEC

Coculture of HUVEC, MSCs and hiPSC-derived hepatic progenitors.

MSC-driven condensation on Matrigel.

• Endothelial networks

• IF: CD31

• RNA-seq analysis

• Protein analysis of the culture supernatants

+

+

Initial % EC: 45%

Surface contact of HE-iPSCs with non-parenchymal cells (HUVECs and MSCs) is required for organoid morphogenesis.

[69]

hPSC-EC HAEC

Coculture of HAEC or iEC, MSCs or iMSC and hiPSC-derived hepatic progenitors.

MSC-driven condensation on Matrigel.

• Endothelial networks

• FACS: CD34, CD31, CD144 at t0

• Proteomic analysis

+

+

Initial % EC: 40%

Differences on the organoid metabolic rate and TGF-β and Wnt signaling pathways were observed depending on the source of EC and MSC used.

[73]

hPSC-EC

Coculture of iEC, iMSCs and hiPSC-derived hepatic progenitors.

iMSC-driven condensation on Matrigel.

• Endothelial networks

• IF: CD144, CD31

• scRNA-seq analysis

• In vivo: Anastomosis with host mouse blood vessels surrounded by pericytes

+

+

Completely hiPSC-derived liver organoids outperformed their counterpart made of HUVEC and MSCs.

Inhibition of KDR decreased the number of endothelial networks and hampered hepatocyte maturation.

[24, 72]

Kidney

hPSC-EC

Co-differentiation of hiPSC into kidney and endothelial lineage and self-assembling on transwells.

• Endothelial networks with lumen

• IF: KDR, CD31, SOX17

–

–

[79, 80]

hPSC-EC HUVEC

Co-differentiation of hiPSC into kidney and endothelial lineage.

Self-assembling in low-attachment plates and further culture on transwells.

• Endothelial networks with lumen

• IF: KDR, CD31, CD34

• scRNA-seq analysis: Different subsets of EC

• In vivo: Perfused and fenestrated vessels

+

–

Maximum 3% of EC at d14

[39]

hPSC-EC

Co-differentiation of human pluripotent epiblast spheroids into kidney and endothelial lineage on Matrigel/Collagen gels.

• Endothelial cords around capsule and tubular structures

• IF: CD31, vWF

–

–

Organoids survived for up to 2 months.

[78]

hPSC-EC, HUVECS, HNDFs, and adult GMECs

Co-differentiation of hiPSC into MM and endothelial lineage in suspension.

From d11 to d14, organoids were culture on perfused chips.

• Endothelial networks with lumen

• IF and FACS: CD31, MCAM and KDR

• mRNA expression: CD31

+

–

[81]

Brain

hPSC-EC

EB co-differentiation of neural progenitors and endothelial cells.

• Vascular-like structures at d30;

• IF: CD144, CD31, vWF, KDR and tight junctions^: OCLN, αZO-1

• EM: Tight junctions

• mRNA expression: CD144, CD31, KDR, TEK, vWF, OCLN, CD34, CLDN5, OCLN, TJP1, ABCB1 and GLUT1

• In vitro perfusion with Dextran-FITC.

• TEER: 351 ± 10 Ω cm^− 2

• scRNA-seq analysis

• In vivo: Anastomosis of human CD31⁺ networks with mouse blood vessels

+

–

EC came from modified hiPSC expressing ETV2 from day 18 of differentiation.

Organoids survived up to 4 months and reached 3,5–4 mm of diameter.

Vascularized organoids displayed better neural differentiation and neural functions.

[85]

hPSC-EC

EB co-differentiation of neural progenitors and endothelial cells in low-attachment plates and further, embedded in Matrigel droplets.

• Tubule-like structures

• IF: CD31, vWF, CLDN5

• mRNA expression: ANGPT1, CD31

• RNA-seq analysis: 106 TJ-related genes and GPR124

+

+

Organoids survived up to 4 months.

α-SMA pericytes surrounded CD31⁺ EC.

[84]

hPSC-EC

Fusion of hPSC-derived cortical and vascular spheroids.

• IF: CD31, CD144, Z0–1

• FACS: CD31

• mRNA expression: BCRP, PGP, GLUT-1

–

–

NPSC: EC: hMSC (1:2:3)

Organoids survived up to 2 months.

[83]

hPSC-EC

Co-culture of d34-brain organoids with ECs in Matrigel.

• Tubular structures

• IF: CD31

• In vivo: human CD31⁺ vessels.

+

+

Organoids survived up to 2 months.

Only in vitro vascularized organoids survived in vivo 2 weeks after transplantation.

[82]